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Technology in Cancer Research &... 2020The biological function of deoxycytidine kinase in tumor is not yet clear, and there are a few studies relating to the correlation of deoxycytidine kinase gene with the...
BACKGROUND
The biological function of deoxycytidine kinase in tumor is not yet clear, and there are a few studies relating to the correlation of deoxycytidine kinase gene with the occurrence and development of liver cancer.
METHODS
The messenger RNA expression of deoxycytidine kinase was analyzed with the use of the UALCAN and GEPIA database. Moreover, we assessed the function of deoxycytidine kinase on clinical prognosis with Kaplan-Meier plotter database. The relationship between deoxycytidine kinase and cancer immune infiltrates was investigated via Tumor Immune Estimation Resource site. Furthermore, Tumor Immune Estimation Resource was also used to evaluate the correlations between the expression of deoxycytidine kinase and gene marker sets of immune infiltrates.
RESULTS
The deoxycytidine kinase messenger RNA level significantly upregulated in patients with liver cancer compared to normal liver samples. Moreover, the increased expression of deoxycytidine kinase messenger RNA was closely associated with reduced overall survival and disease-free survival in all liver cancers. In addition, deoxycytidine kinase expression displayed a strong correlation with infiltrating levels of macrophages, neutrophils, and dendritic cells in liver cancer, and deoxycytidine kinase expression was positively correlated with diverse immune marker sets in liver cancer.
CONCLUSIONS
All the above findings suggested that increased expression of deoxycytidine kinase was significantly related to unfavorable prognosis in patients with liver cancer. And deoxycytidine kinase is correlated with immune infiltrating levels, including those of B cells, macrophages, neutrophils, and dendritic cells in patients with liver cancer. These findings suggest that deoxycytidine kinase can be used as a prognostic biomarker for determining prognosis and immune infiltration in liver cancer. And deoxycytidine kinase is a potential target for liver cancer therapy, and these preliminary findings require further study to determine whether deoxycytidine kinase-targeting reagents might be developed for clinical application in liver cancer.
Topics: Biomarkers, Tumor; Deoxycytidine Kinase; Female; Follow-Up Studies; Humans; Liver Neoplasms; Lymphocytes, Tumor-Infiltrating; Male; Prognosis; Survival Rate
PubMed: 32588770
DOI: 10.1177/1533033820934133 -
Genes Sep 2019Knowledge about synthetic lethality can be applied to enhance the efficacy of anticancer therapies in individual patients harboring genetic alterations in their cancer...
Knowledge about synthetic lethality can be applied to enhance the efficacy of anticancer therapies in individual patients harboring genetic alterations in their cancer that specifically render it vulnerable. We investigated the potential for high-resolution phenomic analysis in yeast to predict such genetic vulnerabilities by systematic, comprehensive, and quantitative assessment of drug-gene interaction for gemcitabine and cytarabine, substrates of deoxycytidine kinase that have similar molecular structures yet distinct antitumor efficacy. Human deoxycytidine kinase (dCK) was conditionally expressed in the genomic library of knockout and knockdown (YKO/KD) strains to globally and quantitatively characterize differential drug-gene interaction for gemcitabine and cytarabine. Pathway enrichment analysis revealed that autophagy, histone modification, chromatin remodeling, and apoptosis-related processes influence gemcitabine specifically, while drug-gene interaction specific to cytarabine was less enriched in gene ontology. Processes having influence over both drugs were DNA repair and integrity checkpoints and vesicle transport and fusion. Non-gene ontology (GO)-enriched genes were also informative. Yeast phenomic and cancer cell line pharmacogenomics data were integrated to identify yeast-human homologs with correlated differential gene expression and drug efficacy, thus providing a unique resource to predict whether differential gene expression observed in cancer genetic profiles are causal in tumor-specific responses to cytotoxic agents.
Topics: Antimetabolites, Antineoplastic; Cytarabine; Deoxycytidine; Deoxycytidine Kinase; Epistasis, Genetic; Gene Ontology; Gene Regulatory Networks; High-Throughput Screening Assays; Humans; Nucleosides; Pharmacogenetics; Phenomics; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins; Gemcitabine
PubMed: 31575041
DOI: 10.3390/genes10100770 -
Proceedings of the National Academy of... Feb 1991Deoxycytidine (dCyd) kinase is required for the phosphorylation of several deoxyribonucleosides and certain nucleoside analogs widely employed as antiviral and...
Deoxycytidine (dCyd) kinase is required for the phosphorylation of several deoxyribonucleosides and certain nucleoside analogs widely employed as antiviral and chemotherapeutic agents. Detailed analysis of this enzyme has been limited, however, by its low abundance and instability. Using oligonucleotides based on primary amino acid sequence derived from purified dCyd kinase, we have screened T-lymphoblast cDNA libraries and identified a cDNA sequence that encodes a 30.5-kDa protein corresponding to the subunit molecular mass of the purified protein. Expression of the cDNA in Escherichia coli results in a 40-fold increase in dCyd kinase activity over control levels. In dCyd kinase-deficient murine L cells, transfection with dCyd kinase cDNA in a mammalian expression vector produces a 400-fold increase over control in dCyd phosphorylating activity. The expressed enzyme has an apparent Km of 1.0 microM for dCyd and is also capable of phosphorylating dAdo and dGuo. Northern blot analysis reveals a single 2.8-kilobase mRNA expressed in T lymphoblasts at 5- to 10-fold higher levels than in B lymphoblasts, and decreased dCyd kinase mRNA levels are present in T-lymphoblast cell lines resistant to arabinofuranosylcytosine and dideoxycytidine. These findings document that this cDNA encodes the T-lymphoblast dCyd kinase responsible for the phosphorylation of dAdo and dGuo as well as dCyd and arabinofuranosylcytosine.
Topics: Amino Acid Sequence; Base Sequence; Blotting, Northern; Cell Line; Cloning, Molecular; DNA, Neoplasm; Deoxycytidine Kinase; Escherichia coli; Gene Expression; Gene Library; Humans; Molecular Sequence Data; Oligonucleotide Probes; RNA, Messenger; RNA, Neoplasm
PubMed: 1996353
DOI: 10.1073/pnas.88.4.1531 -
Nature Communications Aug 2017Leukemia cells rely on two nucleotide biosynthetic pathways, de novo and salvage, to produce dNTPs for DNA replication. Here, using metabolomic, proteomic, and...
Leukemia cells rely on two nucleotide biosynthetic pathways, de novo and salvage, to produce dNTPs for DNA replication. Here, using metabolomic, proteomic, and phosphoproteomic approaches, we show that inhibition of the replication stress sensing kinase ataxia telangiectasia and Rad3-related protein (ATR) reduces the output of both de novo and salvage pathways by regulating the activity of their respective rate-limiting enzymes, ribonucleotide reductase (RNR) and deoxycytidine kinase (dCK), via distinct molecular mechanisms. Quantification of nucleotide biosynthesis in ATR-inhibited acute lymphoblastic leukemia (ALL) cells reveals substantial remaining de novo and salvage activities, and could not eliminate the disease in vivo. However, targeting these remaining activities with RNR and dCK inhibitors triggers lethal replication stress in vitro and long-term disease-free survival in mice with B-ALL, without detectable toxicity. Thus the functional interplay between alternative nucleotide biosynthetic routes and ATR provides therapeutic opportunities in leukemia and potentially other cancers.Leukemic cells depend on the nucleotide synthesis pathway to proliferate. Here the authors use metabolomics and proteomics to show that inhibition of ATR reduced the activity of these pathways thus providing a valuable therapeutic target in leukemia.
Topics: Animals; Ataxia Telangiectasia Mutated Proteins; Biosynthetic Pathways; DNA Replication; Deoxycytidine Kinase; Female; Humans; Mice; Mice, Inbred C57BL; Nucleotides; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Ribonucleotide Reductases
PubMed: 28808226
DOI: 10.1038/s41467-017-00221-3 -
Cancer Science Feb 2004Nucleoside analogues which show antimetabolic activity in cells have been successfully used in the treatment of various tumors. Nucleosides such as... (Review)
Review
Nucleoside analogues which show antimetabolic activity in cells have been successfully used in the treatment of various tumors. Nucleosides such as 1-beta-D-arabinofuranosylcytosine (araC), 6-mercaptopurine, fludarabine and cladribine play an important role in the treatment of leukemias, while gemcitabine, 5-fluorouracil and its prodrugs are used extensively in the treatment of many types of solid tumors. All of these compounds are metabolized similarly to endogenous nucleosides and nucleotides. Active metabolites interfere with the de novo synthesis of nucleosides and nucleotides or inhibit the DNA chain elongation after being incorporated into the DNA strand as terminators. Furthermore, nucleoside antimetabolites incorporated into the DNA strand induce strand-breaks and finally cause apoptosis. Nucleoside antimetabolites target one or more specific enzyme(s). The mode of inhibitory action on the target enzyme is not always similar even among nucleoside antimetabolites which have the same nucleoside base, such as araC and gemcitabine. Although both nucleosides are phosphorylated by deoxycytidine kinase and are also good substrates of cytidine deaminase, only gemcitabine shows antitumor activity against solid tumors. This suggests that differences in the pharmacological activity of these nucleoside antimetabolites may reflect different modes of action on target molecules. The design, in vitro cytotoxicity, in vivo antitumor activity, metabolism and mechanism of action of sugar-modified cytosine nucleosides, such as (2'S)-2'-deoxy-2'-C-methylcytidine (SMDC), 1-(2-deoxy-2-methylene-beta-D-erythro-pentofuranosyl)cytosine (DMDC), 1-(2-C-cyano-2-deoxy-1-beta-D-arabino-pentofuranosyl)cytosine (CNDAC) and 1-(3-C-ethynyl-beta-D-ribo-pentofura-nosyl)cytosine (ECyd), developed by our groups, are discussed here.
Topics: Animals; Antimetabolites, Antineoplastic; Carbohydrates; Cytosine; Humans; Neoplasms; Pyrimidine Nucleosides
PubMed: 14965358
DOI: 10.1111/j.1349-7006.2004.tb03189.x -
The Journal of Biological Chemistry May 2020Nucleoside analogues are a valuable experimental tool. Incorporation of these molecules into newly synthesized DNA ( pulse-labeling) is used to monitor cell...
Nucleoside analogues are a valuable experimental tool. Incorporation of these molecules into newly synthesized DNA ( pulse-labeling) is used to monitor cell proliferation or to isolate nascent DNA. Some of the most common nucleoside analogues used for pulse-labeling of DNA in cells are the deoxypyrimidine analogues 5-ethynyl-2'-deoxyuridine (EdU) and 5-ethynyl-2'-deoxycytidine (EdC). Click chemistry enables conjugation of an azide molecule tagged with a fluorescent dye or biotin to the alkyne of the analog, which can then be used to detect incorporation of EdU and EdC into DNA. The use of EdC is often recommended because of the potential cytotoxicity associated with EdU during longer incubations. Here, by comparing the relative incorporation efficiencies of EdU and EdC during short 30-min pulses, we demonstrate significantly lower incorporation of EdC than of EdU in noninfected human fibroblast cells or in cells infected with either human cytomegalovirus or Kaposi's sarcoma-associated herpesvirus. Interestingly, cells infected with herpes simplex virus type-1 (HSV-1) incorporated EdC and EdU at similar levels during short pulses. Of note, exogenous expression of HSV-1 thymidine kinase increased the incorporation efficiency of EdC. These results highlight the limitations when using substituted pyrimidine analogues in pulse-labeling and suggest that EdU is the preferable nucleoside analogue for short pulse-labeling experiments, resulting in increased recovery and sensitivity for downstream applications. This is an important discovery that may help to better characterize the biochemical properties of different nucleoside analogues with a given kinase, ultimately leading to significant differences in labeling efficiency of nascent DNA.
Topics: Biological Transport; Cell Line; Cytomegalovirus; Deoxycytidine; Deoxyuridine; Fibroblasts; Herpesvirus 1, Human; Herpesvirus 8, Human; Humans; Retinal Pigment Epithelium
PubMed: 32205447
DOI: 10.1074/jbc.RA119.012378 -
Proceedings of the National Academy of... Mar 2010Deoxycytidine kinase (dCK) is a rate-limiting enzyme in deoxyribonucleoside salvage, a metabolic pathway that recycles products of DNA degradation. dCK phosphorylates...
Deoxycytidine kinase (dCK) is a rate-limiting enzyme in deoxyribonucleoside salvage, a metabolic pathway that recycles products of DNA degradation. dCK phosphorylates and therefore activates nucleoside analog prodrugs frequently used in cancer, autoimmunity, and viral infections. In contrast to its well established therapeutic relevance, the biological function of dCK remains enigmatic. Highest levels of dCK expression are found in thymus and bone marrow, indicating a possible role in lymphopoiesis. To test this hypothesis we generated and analyzed dCK knockout (KO) mice. dCK inactivation selectively and profoundly affected T and B cell development. A 90-fold decrease in thymic cellularity was observed in the dCK KO mice relative to wild-type littermates. Lymphocyte numbers in the dCK KO mice were 5- to 13-fold below normal values. The severe impact of dCK inactivation on lymphopoiesis was unexpected given that nucleoside salvage has been thought to play a limited, "fine-tuning" role in regulating deoxyribonucleotide triphosphate pools produced by the de novo pathway. The dCK KO phenotype challenges this view and indicates that, in contrast to the great majority of other somatic cells, normal lymphocyte development critically requires the deoxyribonucleoside salvage pathway.
Topics: Animals; B-Lymphocytes; Deoxycytidine Kinase; Exons; Gene Targeting; Lymphoid Tissue; Lymphopoiesis; Mice; Mice, Knockout; Models, Biological; T-Lymphocytes
PubMed: 20080663
DOI: 10.1073/pnas.0913900107 -
International Journal of Molecular... Apr 2022The ergothioneine transporter ETT (formerly OCTN1; human gene symbol ) is a powerful and highly specific transporter for the uptake of ergothioneine (ET). Recently,...
The ergothioneine transporter ETT (formerly OCTN1; human gene symbol ) is a powerful and highly specific transporter for the uptake of ergothioneine (ET). Recently, Sparreboom et al. reported that the ETT would transport nucleosides and nucleoside analogues such as cytarabine and gemcitabine with the highest efficiency. In our assay system, we could not detect any such transport. Subsequently, Sparreboom suggested that the intracellular metabolization of the nucleosides occurs so fast that the original compounds cannot be detected by LC-MS/MS after inward transport. Our current experiments with 293 cells disprove this hypothesis. Uptake of gemcitabine was easily detected by LC-MS/MS measurements when we expressed the Na/nucleoside cotransporter CNT3 (). Inward transport was 1280 times faster than the intracellular production of gemcitabine triphosphate. The deoxycytidine kinase inhibitor 2-thio-2'-deoxycytidine markedly blocked the production of gemcitabine triphosphate. There was no concomitant surge in intracellular gemcitabine, however. This does not fit the rapid phosphorylation of gemcitabine. Uptake of cytarabine was very slow, but detection by MS was still possible. When the ETT was expressed and incubated with gemcitabine, there was no increase in intracellular gemcitabine triphosphate. We conclude that the ETT does not transport nucleosides.
Topics: Chromatography, Liquid; Cytarabine; Deoxycytidine; Ergothioneine; Humans; Organic Cation Transport Proteins; Tandem Mass Spectrometry; Gemcitabine
PubMed: 35563081
DOI: 10.3390/ijms23094690 -
Journal of Clinical Pathology Jul 2005Deoxycytidine kinase (dCK) is responsible for the activation of several clinically important deoxynucleoside analogues used for the treatment of haematological and solid...
BACKGROUND
Deoxycytidine kinase (dCK) is responsible for the activation of several clinically important deoxynucleoside analogues used for the treatment of haematological and solid malignancies.
AIM
To measure dCK expression in tumour cells from different origins.
METHOD
A rabbit antihuman dCK antibody was used for the immunocytochemical detection of dCK expression in three leukaemic cell lines (HL60, U937, and CCRF-CEM) and 97 patient samples (paediatric acute myeloid leukaemia (AML) and lymphoid leukaemia (ALL), retinoblastoma, paediatric brain tumours, and adult non-small cell lung cancer (NSCLC)).
RESULTS
CCRF-CEM, U937, and HL60 cells stained positively for dCK and the degree of expression correlated with dCK activity. dCK expression varied between tumour types and between individual patients within one tumour type. dCK was located predominantly in the cytoplasm. The staining intensity was scored as negative (0), low (1+), intermediate (2+), or high (3+). Expression of dCK was high in AML blasts. In contrast, brain tumour samples expressed low amounts of dCK. dCK staining ranged from low (1+) to high (3+) in ALL blasts, retinoblastoma, and NSCLC tissue samples. Staining was consistent (interobserver variability, 88%; kappa = 0.83) and specific. Western blotting detected the dCK protein appropriately at 30 kDa, without additional bands.
CONCLUSIONS
Immunocytochemistry is an effective and reliable method for determining the expression of dCK in patient samples and requires little tumour material. This method enables large scale screening of dCK expression in tumour samples.
Topics: Adult; Biomarkers, Tumor; Blotting, Western; Brain Neoplasms; Carcinoma, Non-Small-Cell Lung; Child; Cytoplasm; Deoxycytidine Kinase; Humans; Immunoenzyme Techniques; Leukemia; Lung Neoplasms; Neoplasms; Retinal Neoplasms; Retinoblastoma; Tumor Cells, Cultured
PubMed: 15976334
DOI: 10.1136/jcp.2004.023861 -
International Journal of Gynecological... 2005Gemcitabine (2'2'-difluorodeoxycytidine [dFdC]) is a synthetic analog of deoxycytidine with two fluorine atoms at the 2' position of the carbohydrate. As a hydrophobic... (Review)
Review
Gemcitabine (2'2'-difluorodeoxycytidine [dFdC]) is a synthetic analog of deoxycytidine with two fluorine atoms at the 2' position of the carbohydrate. As a hydrophobic molecule, dFdC competes for intracellular access via membrane-associated nucleoside transporter proteins. Following intracellular transport, dFdC is phosphorylated sequentially by deoxycytidine kinase to gemcitabine triphosphate, which inhibits ribonucleotide metabolism, hinders DNA processing, and increases accumulation of intrastrand adducts and interstrand cross-links, thereby leading to a G1 block in the cell cycle. dFdC monotherapy has been extensively evaluated at doses of 800-1250 mg/m2. dFdC is generally well tolerated, with the most frequently occurring dose-limiting toxicities being hematologic, noncumulative, and easily managed by dose alteration. Several studies involving treatment of recurrent ovarian cancer patients with dFdC monotherapy, most of whom had platinum-resistant disease and/or prior exposure to paclitaxel, led to overall response rates of 14-22% and a median duration of response of 4.0-10.6 months. An additional one third of the participants experienced stable disease for an overall clinical benefit in approximately one half of the treated patients. Tumor cells with a multidrug resistance phenotype have increased sensitivity to dFdC (collateral sensitivity). As dFdC is unaffected by platinum resistance, and not susceptible to classic multidrug resistance, it could be particularly beneficial to administer following treatment with agents that induce multidrug resistance. Integration of dFdC with platinum and/or radiation should also be investigated.
Topics: Antimetabolites, Antineoplastic; DNA, Neoplasm; Deoxycytidine; Drug Resistance, Multiple; Female; Humans; Neoplasm Recurrence, Local; Ovarian Neoplasms; Phenotype; Phosphorylation; Gemcitabine
PubMed: 15839953
DOI: 10.1111/j.1525-1438.2005.15352.x